Non-detection of human herpesvirus 8 (HHV-8) DNA in HHV-8-seropositive blood donors from three Brazilian regions.

Human herpesvirus 8 (HHV-8), also known as Kaposi's sarcoma-associated herpesvirus (KSHV), is the etiologic agent of all forms of Kaposi's sarcoma, primary effusion lymphoma and the plasmablastic cell variant of multicentric Castleman disease. In endemic areas of sub-Saharan Africa, blood transfusions have been associated with a substantial risk of HHV-8 transmission. By contrast, several studies among healthy blood donors from North America have failed to detect HHV-8 DNA in samples of seropositive individuals. In this study, using a real-time PCR assay, we investigated the presence of HHV-8 DNA in whole-blood samples of 803 HHV-8 blood donors from three Brazilian states (São Paulo, Amazon, Bahia) who tested positive for HHV-8 antibodies, in a previous multicenter study. HHV-8 DNA was not detected in any sample. Our findings do not support the introduction of routine HHV-8 screening among healthy blood donors in Brazil. (WC = 140).

Prevalence of, and risk factors for Kaposi's sarcoma-associated herpesvirus infection among blood donors in Brazil: a multi-center serosurvey.

Kaposi's sarcoma-associated herpesvirus (KSHV) is endemic in the Amazon and rare in southern regions of Brazil. However, geographical distribution and epidemiological correlates of infection in this large country are still poorly defined. To estimate the seroprevalence of, and risk factors for, KSHV infection in Brazil, a multi-center study was conducted among 3,493 first-time voluntary unpaid blood donors from Salvador, Sao Paulo and Manaus. Antibodies against KSHV were detected using a whole-virus ELISA validated prior to the serosurvey. Antibodies against the latency-associated nuclear antigen (LANA) were detected by immuno-fluorescence assay (IFA) among ELISA-positive sera and a random sample of ELISA-negative sera. Overall, seroprevalence of KSHV by whole-virus ELISA was 21.7% (95% confidence interval (CI): 20-23.4%) in men and 31.7% (95% CI: 29-34.3%) in women (P<0.0001). KSHV antibodies were detected by IFA-LANA in 3% (95% CI: 2-4.3%) of 867 ELISA-positive samples and in none of 365 randomly selected ELISA-negative samples. In multivariate analysis, KSHV seroprevalence by whole-virus ELISA was independently associated with female sex (odds ratio [OR]=1.6, 95% CI: 1.4-1.9); residence in the Amazon (OR=1.4, 95% CI: 1.2-1.8; compared to Salvador); Caucasian ethnicity (OR=1.3, 95% CI: 1.1-1.6) and herpes simplex virus type 2 (HSV-2) infection (OR=1.3, 95% CI: 1.1-1.6). KSHV seroprevalence did not significantly increase with age, nor was it associated with self-reported sexual behavior. KSHV seroprevalence is high among Brazilian blood donors, particularly from the Amazon region. This study supports the co-existence of sexual and non-sexual routes of KSHV transmission in this population.

Seroreactivity to Kaposi's sarcoma-associated herpesvirus (human herpesvirus 8) latent nuclear antigen in AIDS-associated Kaposi's sarcoma patients depends on CD4+ T-cell count.

In AIDS/Kaposi's sarcoma (KS) patients, the sensitivity of immunofluorescence assays for detecting antibodies against latent nuclear antigen ranges from 52% to 93%. However, in classic and African KS, sensitivities above 90% have been reported systematically. This study evaluates whether CD4+ T-cell count affects seroreactivity to KSHV LANA and to lytic antigens in AIDS/KS patients. Kaposi's sarcoma-associated herpesvirus (KSHV) latent (IFA-LANA) and lytic (IFA-Lytic and ORF65/K8.1 EIA) antibodies were screened in 184 consecutive samples taken from 36 AIDS/KS patients grouped according to their CD4+ counts as follows: <100 (group A), 100-300 (group B), and >300 (group C) cells/mm(3). At enrollment, the immunofluorescence assay for the detection of antibodies against latent nuclear antigen (IFA-LANA) was positive in 3/11(27.2%) group A patients, in 10/11 (90.9%) group B patients, and in 14/14 (100%) group C patients (P < 0.01). Seropositivity to lytic antigens did not differ according to CD4+ T-cell count. Considering IFA-Lytic and ORF65/K8.1 EIA, seropositivity for lytic antigens was 100% in all three patient groups. In patients whose CD4+ count improved during follow-up, IFA-LANA seroconversion occurred; unstable counts resulted in a decrease in LANA antibody titers while the persistence of high counts resulted in unchanged, elevated antibody titers. In conclusion, LANA seroreactivity in AIDS/KS patients, as assessed by an immunofluorescence assay, depends on CD4+ T-cell count, rendering this evaluation important in the interpretation of seroepidemiological studies of KSHV infection in AIDS patients. To evaluate future serological tests based on latency-associated antigens, the selection of sera from KS patients with CD4+ cell count >300 cells/mm(3) as a positive gold standard is recommended.

Sensitivity and specificity of three ELISA-based assays for discriminating primary from secondary acute dengue virus infection.

BACKGROUND: Discrimination between primary and secondary dengue virus infection traditionally has been performed using the hemagglutination inhibition (HI) test. However, this test has practical limitations and disadvantages. OBJECTIVE: To evaluate the ability of three ELISA-based methods (IgG avidity test, IgM/IgG ratio and IgG titer) to discriminate primary from secondary dengue infection. STUDY DESIGN: Serum samples from convalescent-phase patients with confirmed acute, primary (n=46) or secondary (n=33) dengue virus infection were tested using three ELISA-based methods. A ROC curve was employed to establish the cut-off points and to evaluate the ability of the three methods to distinguish between acute, primary and secondary dengue virus infection. RESULTS: All three assays exhibited sensitivity and negative predictive values of 100% for defining secondary infection. The specificity and positive predictive values were respectively 97.8% and 93.7% for the IgG avidity test, 95.7% and 88.2% for the IgM/IgG ratio assays, and 97.8% and 93.7% for the IgG titer assay. CONCLUSION: All three ELISA-based assays proved reliable tools for discriminating between acute, primary and secondary dengue virus infection when using serum samples from convalescent-phase patients.

Evaluation of a commercial real-time PCR kit for detection of dengue virus in samples collected during an outbreak in Goiania, Central Brazil, in 2005.

In the past 2 decades, dengue has reemerged in Brazil as a significant public health problem. Clinicians demand a diagnostic test with high sensitivity that is applicable during the early symptomatic phase. We aimed to test two distinct molecular methods on samples from suspected dengue cases during an outbreak in Central Brazil. Acute-phase serum specimens from 254 patients suspected of having dengue were collected during 2005 in the city of Goiânia, Central Brazil. Samples were blindly evaluated by real-time and multiplex PCR in addition to routine immunoglobulin M serology and virus culture. Overall, acute dengue was confirmed by serology, multiplex PCR, or virus isolation for 80% of patients (203/254). Another four patients presented real-time PCR-positive results as the unique marker of dengue. Higher real-time PCR positivity levels and viral loads were observed in the early symptomatic phase of disease (< or =5 days) than after this period. Multiplex and real-time PCR assays presented a high kappa agreement (0.85). According to multiplex PCR, 60 samples harbored dengue virus type 3 (DEN-3), 4 samples harbored DEN-2, and 1 sample displayed a pattern compatible with a double infection with DEN-2 and -3. The dengue virus real-time kit was found to be practical and adjustable for high throughput, to display the best performance in the early symptomatic phase of dengue cases, and to be valuable for confirming dengue diagnosis in a timely manner.

Comparative study of Kaposi's sarcoma-associated herpesvirus serological assays using clinically and serologically defined reference standards and latent class analysis.

Accurate determination of infection with Kaposi's sarcoma-associated herpesvirus (KSHV) has been hindered by the lack of a "gold standard" for comparison of serological assays used to estimate KSHV prevalence in serosurveys conducted in different settings. We have evaluated the performance of five in-house (developed at University College London [UCL], United Kingdom, and at the virology laboratory of the Instituto de Medicine Tropical [IMT] in Sao Paulo, Brazil) and two commercial (ABI and DIAVIR) serological assays to detect antibodies to latency-associated nuclear antigen (LANA) and to lytic KSHV antigens. We used a variety of serum samples assembled to represent populations likely to be at high, intermediate, and low risk of KSHV infection in Brazil. Composite reference standard panels were prepared based on clinical and serological parameters, against which assay performances were assessed using conventional Bayesian statistics and latent class analysis (LCA). Against the clinical reference standard, in-house immunofluorescence assays to detect anti-LANA antibodies (IFA-LANA) produced at UCL and IMT had similar performances, with sensitivities of 61% (95% confidence interval [CI], 48% to 74%) and 72% (95% CI, 58% to 83%) and specificities of 99% (95% CI, 94% to 100%) and 100% (95% CI, 96% to 100%), respectively, and only the IMT IFA-LANA was included in LCA, together with the IMT IFA-lytic and four enzyme-linked immunosorbent assays (ELISAs). The LCA indicated that the IMT whole-virus ELISA performed best (sensitivity, 87% [95% CI, 81% to 91%]; and specificity, 100% [95% CI, 98% to 100%]), confirming the results obtained with the conventional statistical approach. Commercially available ELISA-based tests yielded the lowest specificities using a spectrum of serum samples. The evaluation of KSHV serological assays is warranted before planning serosurveys in various settings.

Seroprevalence of antibodies against varicella-zoster virus and response to the varicella vaccine in pediatric renal transplant patients.

The severity of varicella-zoster virus (VZV) in immunocompromised children, especially in those receiving renal transplants, is well known. However, the use of live attenuated virus vaccine in this population is controversial. This study aimed to: (i) assess the immunization status of pediatric renal transplant recipients at our center; (ii) determine the anti-VZV antibody titers in such patients; (iii) evaluate the response to VZV vaccine in seronegative children and in those who present low antibody titers (defined as <500 mAU/mL). Vaccinated children were monitored for adverse effects for 8 wk after vaccination. Fifty patients with a mean age of 13.7 yr (range, 3-17 yr) were enrolled. In 49, blood samples were collected and antibodies were screened using ELISA. Seropositivity to VZV was found in 43 (88%), and antibody titers were >/=500 mAU/mL in 37 (75.5%). Of the 12 children who were eligible for vaccination and had antibody titers <500 mAU/mL, one developed varicella before vaccination, two did not meet the inclusion criteria, and three parents refused the vaccination. In the six vaccinated children, there were no adverse reactions to the vaccine, and four (66.6%) responded with anti-VZV titers >/=500 mAU/mL 6-8 wk after vaccination. In conclusion, after renal transplantation, varicella vaccine is safe with a 66% rate of conversion to high antibody titers.

Human herpesvirus 8 (HHV-8) infection in HIV/AIDS patients from Santos, Brazil: seroprevalence and associated factors.

GOAL: The goal of this study was to evaluate the seroprevalence of human herpesvirus 8 (HHV-8) infection among HIV-infected individuals from Brazil and the associated risk factors. STUDY: A cross-sectional survey was carried out with 497 HIV/AIDS outpatients attending the local AIDS Reference Center in Santos (southeastern Brazil) between February 1997 and January 1998 had serum samples screened for anti-HHV-8 antibodies. Patients were considered seropositive whenever reactivity was observed in at least 1 of 3 tests (immunofluorescence assays for latent nuclear and lytic antigens and orf65 recombinant antigen enzyme-linked immunosorbent assay). RESULTS: Overall HHV-8 seroprevalence was 13.9% (95% confidence interval [CI], 10.9-17.6). HHV-8 coinfection was significantly more frequent in men (18.7%; 95% CI, 14.1-23.4) than in women (7.8%; 95% CI, 4.2-11.3) (P < 0.001). According to the mode of HIV acquisition among males, seroprevalence of HHV-8 infection was significantly higher in men who have sex with men when compared with the other groups (32.4% vs. 10.0%, P < 0.001). Multivariate logistic regression revealed HHV-8 infection among men to be independently associated with sexual orientation (adjusted odds ratio [AOR], 5.5 for homosexuals; AOR, 2.8 for bisexuals). No significant risk factor for HHV-8 infection could be demonstrated for HIV-infected women in this cohort, CONCLUSIONS: This study provides further evidence that men who have sex with men are at higher risk of HHV-8 infection and shows that the epidemiologic pattern of this infection among HIV/AIDS patients from Santos, Brazil, is similar to that described in other countries with a low incidence of Kaposi's sarcoma.

Use of an immunoglobulin G avidity test to discriminate between primary and secondary dengue virus infections.

An enzyme-linked immunosorbent assay-based immunoglobulin G (IgG) antibody avidity test was evaluated by using sera from 57 patients with acute dengue infection. Overall, 55 of 57 patients were correctly classified (27 of 27 with primary dengue and 28 of 30 with secondary dengue). We conclude that the IgG avidity test can be useful for differentiating between acute, primary, and secondary dengue infections.

[Prevalence of varicella-zoster virus antibodies in young adults from different Brazilian climatic regions]. / Prevalência de anticorpos para o vírus da varicela-zoster em adultos jovens de diferentes regiões climáticas brasileiras.

To evaluate the prevalence of varicella-zoster virus infection in young adults from different Brazilian urban regions, 975 serum samples from blood donors aged 20 to 29 years, from tropical climate cities (Salvador and Fortaleza) and from temperate climate cities (S o Paulo, Curitiba and Porto Alegre) were tested by an in-house ELISA for detection of anti-varicella-zoster virus IgG antibodies. The overall prevalence was 94.2%. The lowest rate was observed in Fortaleza (88.7%) and the highest in Curitiba (99.5%). Seroprevalence in tropical regions of Brazil (89.4%) was significantly higher than in temperate regions (97.3%), a similar pattern to that observed in other tropical countries.